PROJECT SUMMARY The treatment of cancer has been revolutionized by the development of immune-based therapies designed to boost the number and function of cytotoxic T cells that kill tumor cells. However, it is already apparent that this strategy alone will not benefit all patients, as the majority of cancers generate a highly immunosuppressive tumor microenvironment (TME) that shuts down even the most potent T cells. Furthermore, as a consequence of ramping up the immune system, these immune stimulating drugs instigate significant autoimmune toxicity. This proposal aims to develop a radical new strategy to circumvent both challenges by identifying immunosuppressive regulatory mechanisms unique to cancerous tissues, thereby uncovering pathways active only in tumors that can be targeted to generate precision cancer immunotherapies that preserve immune tolerance in healthy tissues. The focus will be on Regulatory T cells (Tregs), a subset of immunosuppressive CD4+ T cells that lie at the fulcrum of immunity, directing cells of the immune system to go or stop. In cancer, Tregs infiltrate tumors and dampen anti-tumor immune responses, but Tregs also play an essential role in preventing autoimmunity. This proposal will test the hypothesis that human cancers generate an immunosuppressive TME by enforcing a unique epigenetic program in intratumoral Tregs that, if identified, could provide new targets to selectively modulate tumor-infiltrating Tregs and limit autoimmunity. To do this, the research will employ genetically engineered mouse (GEM) cancers, an accurate and flexible platform to test the full complement of factors that make up the TME of human cancers, combined with sophisticated genetic tools to track Treg entry and activity in the TME to resolve: (1) how the native location wherein a cancer arises, (2) how the underlying genetic drivers of cancer, and (3) how the immunogenicity of cancer, shape the epigenetic state of intratumoral Tregs. The identification of epigenetic mechanisms controlling context-specific adaptation of immune cell function in cancer would represent a major breakthrough not only for developing precision cancer immunotherapies, but also for treating infectious diseases and autoimmunity with heightened precision.